A G to A transition at nucleotide 11778 in mitochondrial DNA (mtDNA) in the gene specifying the ND4 subunit of complex I results in an arginine to histidine substitution at amino acid 340. This mtDNA point mutation was linked to Leber Hereditary Optic Neuropathy (LHON), a maternally-inherited human disease that leads to blindness in patients during their 2nd and 3rd decades of life. Since this discovery (Wallace et al., Science 242:1427-1430, 1988), more than 30 other pathogenic point mutations in human polypeptide-coding mtDNA genes have been described. While mtDNA encodes 13 mitochondrial proteins involved in oxidative phosphorylation, the remainder of these proteins are encoded by nuclear DNA, synthesized on cytoplasmic ribosomes, and imported into the mitochondria (usually directed by an N-terminal mitochondrial targeting presequence) (Hartl et al., Science 247:930-938, 1990). Thus, mutations in either mtDNA or nuclear DNA may impair mitochondrial function and thereby result in human disease (Schon EA, Trends Biochem Sci. 25:555-560, 2000).
LHON is the most common of all mitochondrial diseases. Three mtDNA mutations (G3460A, G11778A and T14484C) account for 95% of LHON cases, with the G11778A mutation being the most common, accounting for 50% of LHON cases (Chinnery et al., Ann Neurol. 48:188-193, 2000 and Carelli et al., Ann Neurol. 45:320-328, 1999). Each of the foregoing LHON mutations affects a different subunit of the nicotinamide adenine dinucleotide::ubiquinone oxidoreductase complex (complex I) in the oxidative phosphorylation pathway, where electrons first enter the electron transport chain (Wallace DC, Science, 283:1482-1488, 1999). This large enzyme consists of 7 subunits (ND1, 2, 3, 4, 4L, 5, and 6) encoded by mtDNA, while the remaining 35 subunits are nuclear-encoded (Sazanov et al., Biochemistry 39:7229-7235, 2000). Mitochondrial oxidative phosphorylation deficiency due to mutations in complex I subunit genes is believed to play a pivotal role in development of LHON, although the precise pathophysiologic events precipitating acute visual failure and cellular injury remain incompletely understood. Each LHON mutation alters mtDNA-encoded intrinsic complex I membrane proteins, but surprisingly, results from standard spectrophotometric assays of complex I activity in LHON cells containing the G11778A mutation in the ND4 subunit gene are reduced only slightly (Vergani et al., Biochem Biophys Res Commun. 210:880-888, 1995; Majander et al., FEBS Lett. 292:289-292, 1991; and Larsson NG et al., Ann Neurol. 30:701-708, 1991). Only the G3460A mutation in the ND1 subunit gene reduces complex I activity markedly (Brown et al., J Biol. Chem., 275:39831-39836, 2000 and Cock et al., J Neurol Sci. 165:10-17, 1999). However, clear evidence of complex I deficiency with all three pathogenic mutations comes from polarographic investigations showing impairment of cellular respiration when driven by complex I linked substrates (Majander et al., FEBS Lett. 292:289-292, 1991 and Larsson N G et al., Ann Neurol. 30:701-708, 1991). How these different degrees of changes in complex I function result in the same clinical picture of almost simultaneous bilateral apoplectic visual failure during early adult life is unclear, but reductions in oxidative phosphorylation and cellular injury induced by reactive oxygen species are suspect (Esposito et al., Proc Natl Acad Sci USA 96:4820-4825, 1999 and Brown MD, J Neurol Sci. 165:1-5, 1999).
Unlike most other mitochondrial mutations that impair neurologic and myocardial function and are often fatal, patients with LHON, though blind, have a normal life expectancy. Unfortunately, there is little propensity for spontaneous visual recovery in the G11778A LHON patients, and there is no effective therapy. One of many potential avenues for treatment is to utilize gene therapy to introduce a “normal” gene encoding the defective complex I subunit into the optic nerves of LHON patients. While exogenous genes have been successfully imported into the nuclear genome to protect the optic nerve, (Guy et al., Arch Ophthalmol. 117:929-937, 1999 and Guy J, Proc Natl Acad Sci USA 95:13847-13852, 1998) these methods cannot be applied directly to similarly introduce genes into the mammalian mitochondrial genome.